In view of the decreasing supply of fossil fuels and increasing CO2 emissions, the exploitation of renewable biomass of plant origin as an alternative source of energy is becoming increasingly important. As a result of the continuously increasing and greatly accelerating demand for energy by industrialised and emerging countries, equivalent alternative energy sources need to be found in addition to a more efficient utilisation of fossil energy supplies (Lyco et al., 2009, J. Biotechnol. 142 (1): 78-86).
The focus of research is shifting to the concept of bio-refining where biomass of plant origin is used as a source of both energy and raw materials. The objective of bio-refining is the full use of renewable raw materials for the production of chemicals, fuels and energy. Dry biomass such as straw and wood obtained from raw and waste materials containing lignocellulose is used as the starting material of lignocellulose bio-refining.
Special interest is dedicated to the utilisation of raw materials containing lignocellulose which originate from agriculture and forestry residues such as straw and wood as these are inexpensive and do not compete with the food and feed industry (Lyco et al., 2009, J. Biotechnol. 142 (1): 78-86; Kumar et al., 2008, J. Ind. Microbiol. Biotechnol. 35 (5): 377-391; Peters, 2007, Adv. Biochem. Eng. Biotechnol. 105: 1-30; Kamm et al., 2006, Biochem. Mol. Biol. Int. 44 (2): 283-292). Lignocellulose is the biopolymer which is available in the largest quantities on earth and consists of cellulose, hemicellulose and lignin. The proportion of cellulose is about 30-60%, that of hemicellulose about 20-40% and the lignin proportion is about 10-30%. In contrast to cellulose which is composed of unbranched glucose units, hemicellulose consists of pentoses and hexoses which may have additional carbohydrate branches. Lignin, on the other hand, is a polymer of phenolic molecules (Peters, 2007, Adv. Biochem. Eng. Biotechnol. 105: 1-30).
The use of lignocellulose in bio-refining requires reacting the raw material to obtain useful sugars. For this purpose, lignocellulose is pre-treated by mechanical, thermal and/or chemical methods so as to make the cellulose and hemicellulose more accessible for subsequent hydrolysis (Hendriks & Zeeman, 2009, Bioresour. Technol. 100 (1): 10-18). In enzymatic hydrolysis, especially cellulolytic and xylanolytic enzymes play a critical role. These enzymes which belong to the glycoside hydrolases are capable of decomposing the glycosidic bonds in cellulose and hemicellulose.
In addition to the use of cellulolytic and xylanolytic enzymes of mesophilic micro-organisms, the use of thermostable enzymes is of increasing interest as they are suitable for high-temperature processes which increases both the solubility and hence accessibility of the substrate (Turner et al., 2007, Microb. Cell. Fact. 6: 9). Moreover, thermostable enzymes are characterised by increased specificity of the substrate and stability vis-àvis solvents and detergents (Viikari et al., 2007, Adv. Biochem. Eng. Biotechnol. 108: 121-145; Antranikian G. (2008) in: Industrial relevance of thermophiles and their enzymes—Thermophiles—Biology and Technology at High Temperatures, ed. Robb Fea (CRC Press, Taylor & Francis, Boca Raton), pp 113-160).
Xylan
Xylan, the second-most frequent polysaccharide in nature, is the main component of hemicellulose. It is arranged within the fibril between cellulose and lignin and plays an important role in keeping the micro-fibrils together. Xylans consist of a homopolymer backbone of β-1,4-linked xylopyranose units. This is either linear and unsubstituted, may additionally be acetylated or substituted with arabinosyl and glucuronopyranosyl groups. Therefore, a distinction is made between homoxylan, arabinoxylan, glucuronoxylan and glucuronoarabinoxylan (Saha, 2003, J. Ind. Microbiol. Biotechnol. 30 (5): 279-291; Bergquist et al., 2001, Methods Enzymol 330: 301-319; Kulkarni et al., 1999, FEMS Microbiol. Rev. 23 (4): 411-456). In most cases, xylans are present as complex, highly branched heteropolymers (Collins et al., 2002).
The full hydrolysis of xylans to obtain monosaccharides requires a plurality of enzymes which jointly contribute to degradation (Collins et al., 2005, FEMS Microbiol. Rev. 29 (1): 3-23; Bergquist et al., 2001, Methods Enzymol 330: 301-319). Endoxylanases cleave the glycosidic bonds within the xylan backbone, mainly forming shorter xylol oligosaccharides, but also xylose, xylobiose and xylotriose (Polizeli et al. 2005, Appl. Microbiol. Biotechnol. 67 (5): 577-591; Dwivedi et al., 1996, Appl. Microbiol. Biotechnol. 45 (1-2): 86-93). Xylosidases cleave off xylose monomers from the non-reducing end of xylooligosaccharides and xylobiose, whereas xylan is generally not used as a substrate (Collins et al., 2005, FEMS Microbiol. Rev. 29 (1): 3-23; Polizeli et al., 2005, Appl. Microbiol. Biotechnol. 67 (5): 577-591). Additional enzymes such as α-arabinofuranosidases, α-glucuronidases and acetyl xylan esterases are involved in the release of side groups of heterogeneous xylans. α-Arabinofuranosidases separate arabinose from branched arabinoxylans and arabinans, while α-glucuronidases hydrolyse the α-1,2 bonds between the β-xylopyranosyl backbone and the glucuronic acid. Acetylated xylan is hydrolysed by acetyl xylan esterases which separate the acetyl groups of the xylan (Jaeger et al., 2006, Biokatalyse. Angewandte Mikrobiologie, ed. Antranikian G. Springer Verlag, pp 135-160). Xylanolytic enzymes have been identified both in fungi and bacteria and in archaea.
Glycoside Hydrolases
Glycoside hydrolases are enzymes which hydrolyse the glycosidic bonds between one or more carbohydrates and a residue that does not contain carbohydrates. They cleave a plurality of α- and β-linked substrates and are distinguished in terms of their substrate specificity. Glycoside hydrolases are classified on the basis of similarities of the amino acid sequence or, respectively, percentage identities of the amino acid sequence into so-called glycosidehydrolase families (GH families). In addition to a similar amino acid sequence, the members of a family have a similar three-dimensional structure and the same reaction mechanism (Henrissat, 1991, Biochem. J. 280 (Pt 2): 309-316).
A list of the GH families may be found in the CAZy data base (“Carbohydrate-Active enZymes”) in the Internet (Cantarel et al., 2009). Glycoside hydrolases are divided into two classes according to their reaction mechanism: (1) into enzymes which cleave the glycosidic bonds with reversal of the configuration of the anomeric carbon atom and (2) into enzymes which hydrolyse the glycosidic bonds while maintaining the anomeric configuration (Davies & Henrissat, 1995, Structure 3 (9): 853-859; McCarter & Withers, 1994, Curr. Opin. Struct Biol 4 (6): 885-892). Aspartate and/or glutamate residues have been identified as catalytic amino acids in most glycoside hydrolases; however other amino acid residues may also be involved in cleaving the glycosidic bond (Davies & Henrissat, 1995, Structure 3 (9):853-859).
Glycoside hydrolases consist of a catalytic domain and may contain additional domains binding carbohydrates. This are connected to the catalytic domain by a flexible linker and permit the enzymes to bind to the substrate (Shoseyov et al., 2006; Boraston et al., 2004, Biochem. J. 382 (Pt 3): 769-781). The nomenclature for glycoside hydrolases has been standardised by Henrissat et al. (1998, FEBS Lett. 425 (2): 352-354).
Thus the designation of the enzymes and the genes encoding them is made by indicating the substrate in the form of three letters. The designation for the substrate is followed by the number of the GH family the enzyme belongs to and a letter indicating the order in which the enzymes have been identified. An abbreviation for the species is placed first so as to distinguish similar enzymes of different organisms.
Endoxylanases of Thermophilic Bacteria
A number of endoxylanases of thermophilic bacteria are known which are classified into the glycoside hydrolase families 10, 11 and 43 according to Henrissat et al. (1998, FEBS Lett. 425 (2): 352-354), owing to similarities of the amino acid sequences. Bacteria of the species Caldicellulosiruptor produce endoxylanases with maximum activities at temperatures of 65-70° C. and pH 5.5-6.5. Many endoxylanases are also formed by the anaerobic bacterium Clostridium. 
In addition to the endoxylanase activity, the enzymes XynC and XynX of C. thermocellum display the activity of an endoglucanase (Jung et al., 1998, Biochem. Mol. Biol. Int. 44 (2): 283-292; Hayashi et al., 1997, J. Bacteriol. 179 (13): 4246-4253). The endoxylanases XynC and XynY are known to be located in cellulosomes (Hayashi et al., 1997, J. Bacteriol. 179 (13): 4246-4253; Fontes et al., 1995, Biochem. J. 307 (Pt 1): 151-158).
The most thermo-stable endoxylanases are produced by Thermotoga maritima and T. neapolitana. These show maximum activities at 85-105° C. and half-lives of up to 22 hrs at 90° C. and, respectively, 12 hrs at 95° C. (Zverlov et al., 1996 Appl. Microbiol. Biotechnol. 45 (1-2): 245-247; Saul et al., 1995, Appl. Environ. Microbiol. 61 (11): 4110-4113; Winterhalter & Liebl, 1995, Appl. Environ. Microbiol. 61 (5): 1810-1815). Additional endoxylanases are formed of thermophilic bacteria from the species Geobacillus, Rhodothermus, Thermoanaerobacterium and Thermobifida. 
β-Xylosidases of Thermophilic Bacteria
The β-xylosidases known to date are formed by thermophilic bacteria of the genera Caldicellulosiruptor, Clostridium, Geobacillus, Thermoanaerobacter and Thermoanaerobacterium. They are classified into glycoside hydrolase families 3, 39, 43 and 52 according to Henrissat et al. (1998, FEBS Lett. 425 (2): 352-354) on the basis of similarities of the amino acid sequences. In addition to β-xylosidases, the anaerobic bacterium Clostridium stercorarium also produces endoxylanases and α-arabinofuranosidases and is hence capable of fully degrading arabinoxylan to obtain xylose and arabinan (Adelsberger et al., 2004, Microbiology 150 (Pt 7): 2257-2266).
The gene encoding the β-xylosidase XynB1 of G. stearothermophilus is part of a gene cluster which encodes other enzymes involved in the degradation of xylan. In addition to the gene for the β-xylosidase XynB1, this cluster also contains genes for xylanases and α-glucuronidases (Shulami et al., 1999, J. Bacteriol. 181 (12): 3695-3704). The genes encoding the β-xylosidases of Thermo anaerobacter brockii and Thermoanaerobacterium sp. JW/SL YS485 are also located directly beside genes encoding a β-glucosidase or, respectively, an acetyl xylan esterase (Breves et al., 1997, Appl. Environ. Microbiol. 63 (10): 3902-3910; Lorenz & Wiegel, 1997, J. Bacteriol. 179(17): 5436-5441).
Industrial Applications of Cellulolytic and Xylanolytic Enzymes
The use of cellulolytic and xylanolytic is widespread in the industry. For example, they are used in food and feed production, but cellulases and xylanases are also employed in the paper, pulp and in the textile industry. Their use serves the purpose of increasing yield and improving quality. Moreover, the enzymes are used as an environmentally harmless alternative to conventional methods of treatment.
In the food industry, the cellulolytic and xylanolytic enzymes are used for preparing fruit juices, wine and beer, and in the extraction of oil, preferably olive oil, rapeseed oil and sunflower oil as well as in the baked goods industry. In the preparation of fruit juices, cellulases and hemicellulases are used together with pectinases to increase the yield of the juice and to clarify fruit juices. By treating the flesh of fruit with the enzymes, the formation of juice is enhanced and simultaneously the process time is shortened. By subsequently clarifying the juice with the aid of the enzymes, the viscosity is reduced and the filterability thus improved. However, cellulases and hemicellulases are also used together with pectinases in the preparation of oil, preferably olive oil, rapeseed oil and sunflower oil in order to enhance extraction. These enzymes are also used in the preparation of wines where they contribute to clarification, but also to the extraction of dyes present in the fruit and to improving the aroma of the wine.
Moreover, cellulases are used in breweries to hydrolyse barley glucan so as to facilitate filtration of the beer. Xylanases are also used in the baked goods industry. There, they are used as a flour additive to make the dough easier to process and improve the quality of the baked goods (Beg et al., 2001, Appl. Microbiol. Biotechnol. 56 (3-4): 326-338; Bhat, 2000, Biotechnol. Adv. 18 (5): 355-383; Galante et al., 1998, in: Enzymes, biological control and commercial applications, eds. Harman G E & Kubicek C P, publisher Taylor & Francis, London, Vol. 2, pp. 327-342; Bhat & Bhat, 1997, Biotechnol. Adv. 15 (3-4): 583-620).
Cellulases and xylanases are also used in the feed industry where the enzymes contribute to an increase of the nutritional value and easier digestibility of the feed by digesting the cellulose and hemicellulose in feed products of plant origin (Bhat & Bhat, 1997, Biotechnol. Adv. 15 (3-4): 583-620).
In the paper and pulp industry, cellulases and xylanases are used to modify the wood fibre structure so as to make the pulp easier to process. However, the use of xylanases for bleaching paper is also widespread. In such applications, treatment of the pulp with endoxylanases results in the release of lignin, thus rendering the cell wall of the wood fibres more accessible for bleaching agents, whereby the use of bleaching agents can be significantly reduced (Bhat, 2000, Biotechnol. Adv. 18 (5): 355-383; Buchert et al., 1998, in: Trichoderma & Gliocladium—enzymes, biological control and commercial applications, eds. Harman G E & Kubicek C P, publisher Taylor & Francis, London, Vol. 2, pp 343-363).